Mechanical cell lysis apparatus

ABSTRACT

The present invention relates to a cell lysis apparatus and a manufacturing method thereof, and more particularly, to a cell lysis apparatus, which mechanically performs cell lysis, and a manufacturing method thereof. The cell lysis apparatus includes: an inlet port through which fluid containing cells is supplied; a fluid channel defining a passage through which the cell containing fluid supplied from the inlet port flows; a nano blade array including a plurality of nano blades having pointed leading ends and arranged such that the pointed leading ends of the nano blades are oriented in a direction of the fluid supplied through the inlet port to disrupt the cells passing therethrough; and an outlet port through which the disrupted cells and the fluid are discharged.

TECHNICAL FIELD

The present invention relates to a cell lysis apparatus and amanufacturing method thereof, and more particularly, to a cell lysisapparatus, which mechanically performs cell lysis, and a manufacturingmethod thereof.

BACKGROUND ART

Cell lysis refers to the process of decomposing a cell such that themembrane of the cell is disrupted and the intercellular components areexposed. Cell lysis is primarily performed in cell classification andprotein purification. Cell lysis is generally carried out as an initialstage to separate DNA or RNA in a DNA or RNA amplification process suchas PCR (Polymerase Chain Reaction).

Cell lysis for cell disruption is generally classified into optical,mechanical, acoustic, electrical, and chemical cell lysis.

In optical cell lysis, laser micropulses are radiated to a target cellto form a cavitation bubble such that the cell is disrupted throughexpansion of the cavitation bubble. As such, the optical cell lysis iscarried out by radiating laser micropulses into a particular cell or toan adjacent region thereof, causing deformation of the cell andrequiring a separate laser generator.

In acoustic cell lysis, a cell solution or suspension is placed in achamber within an ultrasound bath, followed by application of ultrasoundwaves to disrupt a target cell. Cell disruption using ultrasound wavesrequires a long time and makes it difficult to obtain uniform energydistribution of the ultrasound waves, thereby providing inconsistentresults.

In electrical cell lysis, an electric field is applied to a cell togenerate a membrane potential for cell disruption. Electrical cell lysisis similar to other methods for cell lysis such as a freezing/thawingmethod, a heating method, an osmotic pressure shock method, and the likein that impact is applied to the cell wall. However, these methods applythermal impact to the cell, causing degradation of cell proteins.

In chemical cell lysis, the cell wall is disrupted using an acid, base,detergent, solvent, chemotropic material, and the like. In particular,generally used is detergent-based cell lysis wherein a detergentdisrupts a lipid double layer of a cell to discharge intercellularcomponents and dissolves membrane proteins. However, such chemical celllysis disadvantageously causes degradation of cell proteins, andrequires separate reagents for cell lysis and removal of the reagentsafter the cell lysis, and a long time for cell lysis.

On the contrary, mechanical cell lysis is carried out using a mechanicalnano structure to disrupt a cell wall. Recently, new lab-on-a chip(LOC)-based cell lysis devices are developed to improve efficiency incell lysis while providing convenience in repeated experimentation.Among various LOC-based cell lysis devices in the art, a mechanicallysis chip minimizes protein degradation, which can occur by heating,electrical impact, or cleansing upon cell lysis.

Carlo et al. suggested a mechanical cell lysis apparatus wherein asilicon substrate is subjected to deep reactive ion etching (DRIB) toform nanoscale scallops on a sidewall in order to disrupt a cellmembrane (D. D. Carlo, K. Jeong and L. P. Lee, “Reagentless mechanicalcell lysis by nanoscale barbs in microchannels for sample preparation”,Lab Chip, 2003, 3, 287-291). However, this apparatus entails high costdue to the silicon DRIE process.

Cell lysates resulting from cell lysis are generally used for proteindetection (such as western blotting), immune precipitation, and thelike. These processes are performed by detecting a certain protein or anintermolecular reaction. It is desirable that cell lysis provide asufficient amount of protein products and a high concentration ofpurified proteins. For this purpose, cell lysis needs to be rapidlyperformed with respect to a target cell using as much fluid as possiblethrough a short passageway, and enables immediate analysis of celllysates.

DISCLOSURE Technical Problem

The present invention is conceived to solve such problems in the relatedart and provides a mechanical cell lysis apparatus, which does notrequire application of an additional product or power to a target cellto minimize cell degradation, has a simple structure, and may bemanufactured at low cost.

In addition, the present invention provides a method of manufacturingthe mechanical cell lysis apparatus.

Technical Solution

In accordance with one aspect, the present invention provides amechanical cell lysis apparatus, which includes: an inlet port throughwhich fluid containing cells is supplied; a fluid channel defining apassage through which the cell containing fluid supplied from the inletport flows; a nano blade array including a plurality of nano bladeshaving pointed leading ends and arranged such that the pointed leadingends of the nano blades are oriented in a direction of the fluidsupplied through the inlet port to disrupt the cells passingtherethrough; and an outlet port through which the disrupted cells andthe fluid are discharged.

In accordance with another aspect, the present invention provides amethod of manufacturing a mechanical cell lysis apparatus, whichincludes an inlet port through which fluid containing cells is supplied;a fluid channel defining a passage through which the cell containingfluid supplied from the inlet port flows; a nano blade array including aplurality of nano blades having pointed leading ends and arranged suchthat the pointed leading ends of the nano blades are oriented in adirection of the fluid supplied through the inlet port to disrupt thecells passing therethrough; and an outlet port through which thedisrupted cells and the fluid are discharged. The method includes:preparing a main panel comprising a silicon substrate; forming anetching mask on an upper side of the silicon substrate; and forming thenano blade array and the fluid channel through crystalline wet etching,with sidewalls of the nano blades arranged in a particular directionwith respect to a particular plane of the silicon substrate, such thatnano blades have pointed leading ends.

Advantageous Effects

According to the present invention, the cell lysis apparatus may achieveefficient disruption of cells without cell degradation caused byapplication of thermal, electrical or chemical impact to the cells.

In addition, the cell lysis apparatus according to the present inventionpermits rapid cell lysis and immediate analysis of cell lysates.Further, the cell lysis apparatus according to the present invention maybe directly connected to a manual type syringe, thereby facilitatingcell lysis.

Furthermore, the cell lysis apparatus according to the present inventionmay be manufactured as a disposable cell lysis apparatus, therebyproviding economic feasibility while reducing contamination due to reuseof the apparatus.

DESCRIPTION OF DRAWINGS

FIG. 1 is a plan view of a cell lysis apparatus in accordance with oneexemplary embodiment of the present invention.

FIG. 2 is an enlarged perspective view of nano blades of the cell lysisapparatus in accordance with the exemplary embodiment of the presentinvention.

FIG. 3 is a diagram showing a process of forming the nano blades of thecell lysis apparatus in accordance with the exemplary embodiment of thepresent invention.

FIG. 4 is a flow diagram of a method of manufacturing a cell lysisapparatus in accordance with one exemplary embodiment of the presentinvention.

FIG. 5(a) is an enlarged perspective view of a nano blade array formedin FIG. 4(c) and FIG. 5(b) is a plan view of a leading end of a nanoblade.

FIG. 6 is an enlarged perspective view of a cell lysis apparatus inaccordance with another exemplary embodiment of the present invention.

FIG. 7 is a flow diagram of a method of manufacturing a cell lysisapparatus in accordance with another exemplary embodiment of the presentinvention.

FIG. 8 is a graph depicting results of cell lysis using a cell lysisapparatus according to the present invention and results of conventionalchemical cell lysis.

BEST MODE

In accordance with one aspect, the present invention provides amechanical cell lysis apparatus including: an inlet port through whichfluid containing cells is supplied; a fluid channel defining a passagethrough which the cell containing fluid supplied from the inlet portflows; a nano blade array including a plurality of nano blades havingpointed leading ends and arranged such that the pointed leading ends ofthe nano blades are oriented in a direction of the fluid suppliedthrough the inlet port to disrupt the cells passing therethrough; and anoutlet port through which the disrupted cells and the fluid aredischarged.

Advantageously, the nano blade array may be disposed parallel to flow ofthe fluid supplied through the inlet port, and may be disposed acrossthe fluid channel, which connects the inlet port to the outlet port.

Advantageously, a micro-filter array may be placed ahead of the nanoblade array to remove foreign matter from the fluid. In one embodiment,the micro-filter array may include at least two rows of micro-filters,and one row of micro-filters near the inlet port may have a greaterdistance between micro-filters than the other row of micro-filters.

The inlet port may be configured to be attached to a connector connectedto an external device. Here, the external device may be a typicalsyringe.

The inlet port, the fluid channel, the nano blade array, and the outletport may be formed on a main panel by crystalline wet etching. In someembodiments, the inlet port, the fluid channel, the nano blade array,and the outlet port may be formed by injection using stamperscorresponding thereto.

In accordance with one aspect, the present invention provides a methodof manufacturing a mechanical cell lysis apparatus, which includes aninlet port through which fluid containing cells is supplied; a fluidchannel defining a passageway through which the cell containing fluidsupplied from the inlet port flows; a nano blade array including aplurality of nano blades having pointed leading ends and arranged suchthat the pointed leading ends of the nano blades are oriented in adirection of the fluid supplied through the inlet port to disrupt thecells passing therethrough; and an outlet port through which thedisrupted cells and the fluid are discharged. The method includes:preparing a main panel comprising a silicon substrate; forming anetching mask on an upper side of the silicon substrate; and forming thenano blade array and the fluid channel through crystalline wet etching,with sidewalls of the nano blade array arranged in a particulardirection with respect to a particular plane of the silicon substrate,such that the nano blades have pointed leading ends.

The method may further include attaching a cover panel to an upper sideof the etched silicon substrate.

The method may further include separating a plurality of cell lysisapparatuses from each other formed on the main panel.

MODE FOR INVENTION

Exemplary embodiments of the present invention will now be described indetail with reference to the accompanying drawings. It should be notedthat like components will be denoted by like reference numeralsthroughout the specification and the drawings. In addition, descriptionof details apparent to those skilled in the art will be omitted forclarity. Further, it should be understood that the present invention isnot limited to the following embodiments and may be embodied indifferent ways by those skilled in the art without departing from thescope of the present invention.

FIG. 1 is a plan view of a cell lysis apparatus in accordance with oneexemplary embodiment of the present invention, and FIG. 2 is an enlargedperspective view of nano blades of the cell lysis apparatus inaccordance with the exemplary embodiment of the present invention.

The cell lysis apparatus 10 according to one exemplary embodimentincludes an inlet port 12, an outlet port 18, a fluid channel 14, andnano blades 16. In this embodiment, the inlet port 12, the outlet port18, the fluid channel 14 and the nano blades 16 are formed on a mainpanel and a cover may be attached to an upper side of the main panel toconstitute the cell lysis apparatus.

A fluid containing target cells is supplied into the cell lysisapparatus through the inlet port 12. The fluid may be a buffer solution.The cell containing fluid passing through the inlet port 12 is suppliedto the nano blades 16, in which the cell walls are disrupted by the nanoblades 16, and is then discharged through the outlet port 18.

The fluid channel 14 defines a passageway between the inlet port 12 andthe outlet port 18 and is illustrated as being formed to have right andleft walls having a predetermined length in FIG. 1.

A plurality of nano blades 16 is arranged in an array. In someembodiments, the nano blades 16 are arranged parallel to flow of thefluid in the fluid channel 14 and placed substantially in a centralregion of the fluid channel 14. Each of the nano blades 16 is processedto have a pointed leading end and the distance between the nano blades16 is set to be less than the diameter of a cell supplied through theinlet port 12. In some embodiments, the distance between the nano blades16 is set in the range of 1 to 5 μm. In some embodiments, each of thenano blades 16 may have a length (l) of 50 μm, a height (h) of 5010 μm,and a width (w) of 2 μm.

The cell containing fluid is supplied to the inlet port 12 under acertain pressure such that the cells are forced to collide with thepointed leading ends of the nano blades 16 by the fluid pressure appliedto the inlet port 12 and are disrupted by the pointed leading ends ofthe nano blades 16. To this end, the inlet port 12 may be connected to amotor driven pump or to a syringe having a manual type piston such thatthe cell containing fluid can be supplied to the inlet port by the pumpor piston.

Next, a process of forming the nano blades on the main panel will bedescribed.

FIG. 3 is a diagram showing a process of forming the nano blades of thecell lysis apparatus in accordance with the exemplary embodiment.Specifically, FIG. 3 shows a process of forming nano blades byundercutting through wet etching.

The nano blades 16 and the fluid channel 14 are arranged parallel to a[111] crystal plane 22 of a [110] silicon crystal 20. This arrangementallows crystalline wet etching of the silicon crystal 20 without siliconDRIB. During crystalline wet etching of the [110] silicon crystal 20, aninitial shape of flat nano blades is changed to the shape of very sharpnano blades by undercutting convex corners of the [110] silicon crystal20 under a SiO₂ mask, as shown in FIG. 3. Next, this process will bedescribed in more detail. Referring to FIG. 3(a), an etching mask 24having a pattern of plural narrow rectangles is formed on an uppersurface of the [110] silicon crystal 20 to be arranged parallel to the[111] crystal plane 22. Since the rectangles of the etching mask 24 arearranged to have long sides parallel to the [111] crystal plane 22 ofthe [110] silicon crystal 20, the silicon crystal under the long sidesof the etching masks 24 is etched downwards without undercutting. On thecontrary, the short sides of the rectangles of the etching masks 24 arenot parallel to the [111] crystal plane and thus are subjected to severeundercutting. In addition, the convex corners of the rectangle patternare chemically unstable and affected by an etching solution. As etchingcontinues, both undercut sides meet each other at the convex corner andthe pattern of narrow rectangles finally becomes a pattern of very sharpvertical silicon slits, which constitute the nano blades 16. FIGS. 3(b)and 3(c) are a perspective view and a plan view of the final nano blades16 after etching.

The degree of sharpness of the pointed leading ends of the nano blades16 varies depending on etching conditions such as temperature orconcentration of the etching solution. Meanwhile, advantageously, theconvex corners of the rectangle pattern of the etching mask 24 may berounded to facilitate formation of the nano blades 16 through etching.That is, the rounded convex corners of the rectangle pattern of theetching mask 24 allow the leading ends of the nano blades 16 to beprocessed more sharply.

Next, a method of manufacturing a cell lysis apparatus according to oneexemplary embodiment including the process of forming nano bladesthrough wet etching and the structure of the cell lysis apparatus willbe described in more detail.

FIG. 4 is a flow diagram of a method of manufacturing a cell lysisapparatus in accordance with one exemplary embodiment of the presentinvention

First, referring to FIG. 4(a), a silicon-on-glass (SOG) substrate 30 isprepared as a main panel. The SOG substrate 30 may be prepared byjoining a [110] silicon crystal 20 to an upper side of a glass substrate32, followed by processing the [110] silicon crystal 20 to apredetermined thickness (for example, 10 μm) via chemical mechanicalpolishing.

Next, to perform crystalline wet etching of the [110] silicon crystal20, an etching mask 34 is deposited on an upper side of the SOGsubstrate 30, as shown in FIG. 4(b). The etching mask 34 may be composedof metal layers of gold Au and chromium Cr. In some embodiments, the Aulayer has a thickness of 300 nm and the Cr layer has a thickness of 50nm. Sidewalls of nano blades 16 and a fluid channel 14 are arrangedparallel to a [111] crystal plane 22 of the [110] silicon crystal 20.

Regions exposed through the etching mask to form the nano blades 16 andthe fluid channel 14 are subjected to anisotropic etching using anetching solution to have a shape as shown in FIG. 4(c). Here, as theetching solution, a tetramethyl ammonium hydroxide (TMAH) or KOHsolution may be used. During etching, portions for an inlet port 12 andan outlet port 18 may be formed together on an area of the siliconcrystal 20 of the SOG substrate 30.

Finally, as shown in FIG. 4(d), an upper cover 40 is attached to theupper side of the SOG substrate 30 on which the fluid channel 14 and thenano blades 16 are formed through etching. The upper cover 40 may beprovided with an inlet tube 42 and an outlet tube 44 for forming theinlet port 12 and the outlet port 18, respectively. The upper cover 40may be made of a transparent material so as to allow observation of aseries of processes by which cells are supplied through the inlet port12 and then disrupted by the nano blades.

Then, when a cell containing fluid is supplied through the inlet tube 42of the cell lysis apparatus, cells are disrupted by the pointed leadingends of the nano blades 16 while colliding therewith and theintercellular components of the disrupted cells are discharged throughthe outlet tube 44.

FIG. 5(a) is an enlarged perspective view of the nano blade array formedin FIG. 4(c) and FIG. 5(b) is a plan view of a leading end of a nanoblade. As shown in FIGS. 5 (a) and (b), it can be seen that the nanoblade array is efficiently formed through etching as described above.

The cell lysis apparatus according to the embodiment includes the nanoblade array such that cells can be mechanically disrupted thereby. Next,a cell lysis apparatus according to another exemplary embodiment will bedescried.

FIG. 6 is an enlarged perspective view of a cell lysis apparatus inaccordance with another exemplary embodiment of the present invention.

The cell lysis apparatus 50 according to this embodiment includes aninlet port 52, an array of micro-filters 62, an array of nano blades 56,an outlet port 58, and a fluid channel 54 defining a flow passageway. Inthis embodiment, the inlet port 52, the micro-filter array 62, the nanoblades 56, and the outlet port 58 are formed on a main panel 60, and acover panel 70 may be attached to an upper side of the main panel 60.Alternatively, the inlet port 52, the micro-filter array 62, the nanoblades 56 and the outlet port 58 are formed on both an upper surface ofthe main panel 60 and a lower surface of the cover panel 70, followed bycoupling the main panel 60 to the cover panel 70 to constitute the celllysis apparatus.

A fluid containing target cells is supplied into the cell lysisapparatus through the inlet port 52, which is connected at a lowerportion thereof with the fluid channel 54. In some embodiments, theinlet port 52 may be connected at an upper portion thereof to aconnector 72, which connects an outlet of a typical syringe to the inletport 52. When the cell lysis apparatus 50 is connected to the syringe 74through the connector 72, the cell containing fluid may be supplied tothe inlet port 52 by manually operating the piston of the syringe 72. Insome embodiments, the inlet port 52 and the outlet port 58 may be formedcoplanar with the fluid channel 54.

The micro-filter array 62 serves to remove foreign matter such as dustfrom the cell containing fluid. The micro-filter array 62 may becomposed of pillar-shaped filters arranged at a constant distance, whichis greater than the diameter of a target cell for cell lysis. Referringto FIG. 6, the micro-filter array 62 may be a combination of a primarymicro-filter array 62 a and a secondary micro-filter array 62 b. In someembodiments, the primary micro-filter array 62 a may includemicro-filter structures arranged a distance of 100 μm, and the secondarymicro-filter array 62 b may include includes micro-filter structuresarranged a distance of 30 μm.

The nano blades 56 have the same structure as that of the nano bladesdescribed in FIG. 1 to FIG. 5. In this embodiment, for more efficientcell lysis, a plurality of nano blades 56 is arranged in two or morerows as shown in FIG. 6. FIG. 6 shows three rows of nano blades 56.

The outlet port 58 discharges the cells disrupted by the nano blades 56and may be connected to a cell analysis device or to a container forstoring the disrupted cells.

Referring to FIG. 6, the fluid channel 54 is illustrated as generallyhaving a hexagonal shape wherein the fluid channel 54 is enlarged fromone end of the inlet port 52 and is narrowed at a proximal end of theoutlet port 58. However, it should be understood that the shape of thefluid channel 54 is not limited to the shape shown in FIG. 6.

The fluid channel 54 may be provided with one or more flow distributionstructures 64 a, 64 b, 64 c, 64 d to allow the cell containing fluidsupplied through the inlet port 52 to be uniformly distributed withinthe fluid channel 54 before the cell containing fluid is supplied to themicro-filter array 62 and the nano blades 56 via the fluid channel 54. Afirst flow distribution structure 64 a is placed near the one end of theinlet port 52 and allows the fluid supplied through the inlet port 52 tobe efficiently supplied to the micro-filter array 62 through the fluidchannel 54, which has an increasing width from the inlet port 52. Secondand third flow distribution structures 64 b, 64 c are disposed togetherwith the nano blades 56 and serve to distribute the fluid towards rightand left sides when supplied to the nano blades 56 while increasing theflow speed of the fluid such that the membranes of the cells can beefficiently disrupted by the nano blades 56. Furthermore, a fourth flowdistribution structure 64 d is placed downstream of the nano blades 56and serves to allow efficient fluid delivery.

FIG. 7 is a flow diagram of a method of manufacturing the cell lysisapparatus shown in FIG. 6.

First, a thermal silicon oxide layer 76 is placed as an etching mask onan upper side of the main panel 60 having a [110] silicon substratethereon (FIG. 7(a)). Then, a pattern 78 of cell lysis apparatuses isformed on the silicon oxide layer 76 attached to the overall surface ofthe [110] silicon substrate of the main panel 60 (FIG. 7(b)). Etching isperformed using an etching solution to form an inlet port 52, amicro-filter array 62, nano blades 56, an outlet port 58 and a fluidchannel 54, followed by removal of the silicon oxide layer 76 (FIG.7(c)). A cover panel 70 is attached to the upper side of the main panel60 (FIG. 7(d)). Through these operations, a plurality of cell lysisapparatuses 50 may be formed on a single silicon substrate and separatedfrom each other by through-hole etching 80 or the like.

In FIG. 4 and FIG. 7, the structures forming the cell lysis apparatus 50on the main panel 60 are illustrated as being formed by etching.Alternatively, however, the cell lysis apparatus 50 according to thepresent invention may be formed by injection using stamperscorresponding to the inlet port 52, the fluid channel 54, the nanoblades 56, and the outlet port 58.

FIG. 8 is a graph depicting results of cell lysis using a cell lysisapparatus according to the present invention and results of conventionalchemical cell lysis.

For chemical cell lysis, 5×10⁶ cells contained in 500 μl of a PBS bufferwere dissolved in an ice-containing RIPA buffer (50 mM Tris-HCl, pH 8,150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, 2 mM EDTA)for 30 minutes. The mixture of the cells and the lysis agent was stirredeach 5 minutes. In order to remove cell debris, the cell lysates werecentrifuged at 4° C. and at 13,000 rpm for 10 minutes, and supernatantswere used for quantitative analysis of proteins. The proteinconcentrations of the cell lysates by the chemical cell lysis and thecell lysis apparatus according to the invention were determined byBradford assay.

As can be seen from FIG. 8, the cell lysis apparatus according to thepresent invention provides a higher concentration of proteins thanconventional chemical cell lysis. Furthermore, the chemical cell lysisneeded 30 minutes or more, whereas cell lysis using the apparatusaccording to the present invention needed a short period of time of 2minutes or less.

Although some exemplary embodiments have been described herein, itshould be understood by those skilled in the art that these embodimentsare given by way of illustration only, and that various modifications,variations and alterations can be made without departing from the spiritand scope of the invention. Therefore, the embodiments and theaccompanying drawings should not be construed as limiting the technicalspirit of the present invention, but should be construed as illustratingthe technical spirit of the present invention. The scope of theinvention should be interpreted according to the following appendedclaims as covering all modifications or variations derived from theappended claims and equivalents thereof.

The invention claimed is:
 1. A mechanical cell lysis apparatuscomprising: an inlet port through which fluid is supplied, wherein thefluid contains cells; an outlet port through which disrupted cells andthe fluid are discharged; a fluid channel defining a passageway betweenthe inlet port and the outlet port through which the fluid supplied fromthe inlet port flows, wherein the fluid channel has a first end and asecond end, wherein the first end is enlarged from one end of the inletport, the first end forming an acute angle, and the second end isnarrowed at a proximal end of the outlet port; a flow distributionstructure within the fluid channel allowing uniform distribution of thefluid within the fluid channel; a micro-filter array to remove foreignmatter from the fluid, wherein the micro-filter array is downstream ofthe flow distribution structure; and a nano blade array including aplurality of nano blades having pointed leading ends and arranged suchthat the pointed leading ends of the nano blades are oriented in adirection of the fluid supplied through the inlet port to disrupt thecells passing therethrough, wherein the nano blade array is downstreamof the micro-filter array.
 2. The apparatus according to claim 1,wherein the plurality of nano blades in the nano blade array is disposedparallel to flow of the fluid supplied through the inlet port.
 3. Theapparatus according to claim 1, wherein the nano blade array is disposedacross the fluid channel connecting the inlet port to the outlet port.4. The apparatus according to claim 1, wherein the micro-filter arraycomprises at least two rows of micro-filters, one row of micro-filtersnear the inlet port having a greater distance between micro-filters thanthe other row of micro-filters.
 5. The apparatus according to claim 1,wherein the inlet port, the nano blade array, the fluid channel and theoutlet port are formed on a main panel, and a cover panel is attached toan upper side of the main panel.
 6. The apparatus according to claim 5,wherein the inlet port and the outlet port are placed coplanar with thefluid channel connecting the inlet port to the outlet port.
 7. Theapparatus according to claim 5, wherein the cover panel is provided withan inlet tube connected to the inlet port and an outlet tube connectedto the outlet port.
 8. The apparatus according to claim 5, wherein theinlet port is configured to be attached to a connector connected to anexternal device.
 9. The apparatus according to claim 8, wherein theexternal device comprises a syringe.
 10. The apparatus according toclaim 1, wherein the inlet port, the fluid channel, the nano bladearray, and the outlet port are formed on a main panel throughcrystalline wet etching.
 11. The apparatus according to claim 1, whereinthe inlet port, the fluid channel, the nano blade array, and the outletport are formed by injection using stampers corresponding thereto.